Runx3/Pebp2alphaC null mouse gastric mucosa exhibits hyperplasias due to stimulated proliferation and suppressed apoptosis in epithelial cells, and the cells are resistant to growth-inhibitory and apoptosis-inducing action of TGF-beta, indicating that Runx3 is a major growth regulator of gastric epithelial cells. Between 45% and 60% of human gastric cancer cells do not significantly express RUNX3 due to hemizygous deletion and hypermethylation of the RUNX3 promoter region. Tumorigenicity of human gastric cancer cell lines in nude mice was inversely related to their level of RUNX3 expression, and a mutation (R122C) occurring within the conserved Runt domain abolished the tumor-suppressive effect of RUNX3, suggesting that a lack of RUNX3 function is causally related to the genesis and progression of human gastric cancer.

The vacuolating cytotoxin VacA produced by Helicobacter pylori causes massive cellular vacuolation in vitro and gastric tissue damage in vivo, leading to gastric ulcers, when administered intragastrically. Here we report that mice deficient in protein tyrosine phosphatase receptor type Z (Ptprz, also called PTP-zeta or RPTP-beta, encoded by Ptprz) do not show mucosal damage by VacA, although VacA is incorporated into the gastric epithelial cells to the same extent as in wild-type mice. Primary cultures of gastric epithelial cells from Ptprz+/+ and Ptprz-/- mice also showed similar incorporation of VacA, cellular vacuolation and reduction in cellular proliferation, but only Ptprz+/+ cells showed marked detachment from a reconstituted basement membrane 24 h after treatment with VacA. VacA bound to Ptprz, and the levels of tyrosine phosphorylation of the G protein-coupled receptor kinase-interactor 1 (Git1), a Ptprz substrate, were higher after treatment with VacA, indicating that VacA behaves as a ligand for Ptprz. Furthermore, pleiotrophin (PTN), an endogenous ligand of Ptprz, also induced gastritis specifically in Ptprz+/+ mice when administered orally. Taken together, these data indicate that erroneous Ptprz signaling induces gastric ulcers.

Genes involved in the transforming growth factor ␤ (TGF-␤) signaling pathway are frequently altered in several types of cancers, and a gastric tumor suppressor RUNX3 appears to be an integral component of this pathway. We reported previously that apoptosis is notably reduced in Runx3 ؊/؊ gastric epithelial cells. In the present study, we show that a proapoptotic gene Bim was transcriptionally activated by RUNX3 in the gastric cancer cell lines SNU16 and SNU719 treated with TGF-␤. The human Bim promoter contains RUNX sites, which are required for its activation. Furthermore, a dominant negative form of RUNX3 comprised of amino acids 1 to 187 increased tumorigenicity of SNU16 by inhibiting Bim expression. In Runx3 ؊/؊ mouse gastric epithelium, Bim was down-regulated, and apoptosis was reduced to the same extent as that in Bim ؊/؊ gastric epithelium. We confirmed comparable expression of TGF-␤1 and TGF-␤ receptors between wild-type and Runx3 ؊/؊ gastric epithelia and reduction of Bim in TGF-␤1 ؊/؊ stomach. These results demonstrate that RUNX3 is responsible for transcriptional up-regulation of Bim in TGF-␤-induced apoptosis.

In intestinal epithelial cells, inactivation of APC, a key regulator of the Wnt pathway, activates beta-catenin to initiate tumorigenesis. However, other alterations may be involved in intestinal tumorigenesis. Here we found that RUNX3, a gastric tumor suppressor, forms a ternary complex with beta-catenin/TCF4 and attenuates Wnt signaling activity. A significant fraction of human sporadic colorectal adenomas and Runx3(+/-) mouse intestinal adenomas showed inactivation of RUNX3 without apparent beta-catenin accumulation, indicating that RUNX3 inactivation independently induces intestinal adenomas. In human colon cancers, RUNX3 is frequently inactivated with concomitant beta-catenin accumulation, suggesting that adenomas induced by inactivation of RUNX3 may progress to malignancy. Taken together, these data demonstrate that RUNX3 functions as a tumor suppressor by attenuating Wnt signaling.

This mouse line is the first genetically engineered mouse model of DGC and is very useful for clarifying the mechanism underlying gastric carcinogenesis, and provides a new approach to the treatment and prevention of DGC.

To clarify the role of micro (mi) RNAs in gastric carcinogenesis, we studied the expression and function of miRNAs in gastric carcinoma (GC) cells. Initially, we performed microarray analysis using total RNA from 3 human GC cell lines and noncancerous gastric tissue. Among the downregulated miRNAs in GC cells, miR-212 expression was decreased in all 8 GC cell lines examined and a significant decrease of miR-212 expression in human primary GC tissues was also observed in 6 of 11 cases. Transfection of the precursor miR-212 molecule induced decreased growth of 3 GC cell lines. Using 3 different databases, methyl-CpG-binding protein MeCP2 was postulated to be a target of miR-212. As seen on reporter assaying, miR-212 repressed the construct with the MECP2 3 0 -UTR. Ectopic expression of miR-212 repressed expression of the MeCP2 protein but not the MECP2 mRNA level. These data suggest that downregulation of miR-212 may be related to gastric carcinogenesis through its target genes, such as MECP2.

Due to a mistake made during manuscript preparation, the Supplemental Data for this article listed incorrect sequences for the primer sets for methylation-specific PCR (MSP). The correct sequences are given below. These corrections do not affect the findings or conclusions of the study.Primer sets used for detection of methylated DNA were 5 0 -ataaagagaaattaggcgc-3 0 and 5 0 -ataaccctcgaaaaacgcg-3 0 (M3), 5 0 -gatgttt gtttaggtcgtagcggtc-3 0 and 5 0 -ccaaactcgaaattcgccgta-3 0 (M2), and 5 0 -tgcgattggttgcgtttcgc-3 0 and 5 0 -cgaaaatacgcataccgcg-3 0 (M1). Primer sets used for detection of unmethylated DNA were 5 0 -ataaagagaaattaggtgt-3 0 and 5 0 -ataaccctcaaaaaacaca-3 0 (M3), 5 0 -tgtttg tttaggttgtagtggttgt-3 0 and 5 0 -cccccaaactcaaaattcaccata-3 0 (M2), and 5 0 -tgtgattggttgtgttttgt-3 0 and 5 0 -caaaaatacacataccaca-3 0 (M1).

Transforming growth factor ␤ (TGF-␤) has been implicated in the maintenance of homeostasis in various organs, including the gastric epithelium. In particular, TGF-␤-induced signaling was shown to be required for the differentiation-associated physiological apoptosis of gastric epithelial cells, but its mechanism has not been well understood. In this study, the molecular mechanism of TGF-␤-induced apoptosis was analyzed in a human gastric epithelial cell line, SNU16, as an in vitro model. Expression of Smad7 and Bcl-X L , but not viral FLIP, was shown to prevent TGF-␤-induced apoptosis, indicating an exclusive requirement of the activation of Smad signaling pathway and mitochondrial dysfunction followed by activation of caspase-9. In addition, treatment with TGF-␤ induced binding of Bim, a proapoptotic Bcl-2 homology domain 3 (BH3)-only protein, to Bcl-X L , which is dependent on the activation of Smad, and reduction in the expression of Bim by RNA interference decreased the sensitivity to TGF-␤-induced apoptosis. Moreover, we found abnormalities in the gastric epithelium of both Bim and caspase-9 knockout mice; these abnormalities were associated with a defect of physiological apoptosis in gastric epithelial cells. These results indicate for the first time that TGF-␤ is involved in the physiological loss of gastric epithelial cells by activating apoptosis mediated by Smad, Bim, and caspase-9.Gastric epithelial cells exhibit a rapid rate of turnover, which requires an appropriate balance between the proliferation of progenitor cells and the loss of mature cells (12). Loss of mature epithelial cells at the gastric surface is thought to be mediated mainly by physiological cell death, namely, apoptosis, which always occurs at the surface of the gastrointestinal tract. In the gastric mucosa, 1 to 3% of the epithelial cells were reported to show morphological features of apoptosis at any given time under physiological conditions (47). The physiological loss of gastric epithelial cells is necessary for the maintenance of tissue homeostasis associated with the exchange of mature epithelial cells to fresh proliferating cells, and any defects in this epithelial cell death pathway may be a contributing factor to disease development. However, little direct evidence linking the gastric epithelial cell death and apoptosisrelated molecules has been obtained and the molecular regulation of apoptosis of gastric epithelial cells largely remains unclear. In this study, we focused on transforming growth factor ␤ (TGF-␤) as one of the key regulators in this physiological cell death.TGF-␤ is a multifunctional cytokine, which has essential roles in a variety of physiological or pathological processes (9, 42). While TGF-␤ functions as a potent suppressor of cell proliferation, it can also induce or suppress apoptosis in certain types of cells (39,42). Previous studies have established that the acquisition of resistance to TGF-␤ is a critical step for carcinogenesis in many organs (8). Abnormal expression of TGF-␤ receptors and inactive...